Acute myeloid leukemia (AML), the most common form of adult acute leukemia usually occurs around age 60 has a very poor prognosis; most patients survive less than 18 months following diagnosis. AML is a heterogeneous disease with multiple signaling pathways contributing to its pathogenesis. Internal tandem duplications in FMS-like tyrosine kinase receptor 3 (FLT3-ITD) are seen in ~30% of AML patients, and are associated with poor prognosis. Gain-of-function mutations in receptor tyrosine kinase KIT, including mutation at codon 816 (codon 814 in mouse) are also associated with human AML. Despite substantial efforts devoted to the development of FLT3 or KIT inhibitors, the effectiveness of these agents in AML has been limited. There is a critical need to identify effector molecules promoting FLT3-ITD or KITD816V-mediated oncogenic signaling in leukemic cells, which can provide the basis for development of novel therapeutic approaches for AML. The phosphatase of regenerating liver (PRL) family of phosphatases consists of PRL1, PRL2, and PRL3, and represents an intriguing group of proteins currently being validated as biomarkers and therapeutic targets in human cancer. We found that PRL2 is important for hematopoietic stem and progenitor cell proliferation through its role in regulating SCF/KIT signaling (Kobayashi et al., Stem Cells, 2014), but the role of PRL2 in leukemogenesis remains unknown. We recently found that loss of Prl2 decreased the ability of oncogenic KITD814V and FLT3-ITD to promote mouse HSPC proliferation in vitro. Furthermore, we discovered that PRL2 was overexpressed in human AML cell lines, and we found that inhibiting PRL2 activity with a PRL inhibitor (PRLi) decreased the proliferation and survival of AML cell lines and primary AML cells. Based on these preliminary data, we hypothesize that PRL2 is a mediator of oncogenic cytokine signaling in AML cells, and that pharmacologic inhibition of PRL2 will impair leukemic cell viability, and increase effectiveness of leukemia treatment. To test our hypothesis, we propose the following two Specific Aims: Aim 1) Determine the effect of genetic or pharmacologic inhibition of PRL2 on AML cells and Aim 2) Identify the mechanism by which PRL2 contributes to the pathogenesis of AML. Mutations in FLT3 and KIT are found in approximately 40% of AML patients. We found that PRL2 is important for mediating these oncogenic signaling pathways in AML. Therefore, successful completion of these studies are likely to have an important positive impact on healthcare for the citizens of the United States, because establishing PRL2 as a therapeutic target in AML will facilitate the development of PRL2 inhibitors for treating AML, and may be applicable to the treatment of other human cancers, as well.